Abstract
Objectives:
Oral nicotine pouches (ONPs) are increasing in use, yet data on potential oral health effects are scarce. We evaluated whether ONP use is associated with clinical periodontal measures and salivary microbiome dysbiosis in a field-based regional sample. We recruited firefighters, as this population has historically featured an elevated prevalence of oral tobacco use.
Methods:
In this preliminary, cross-sectional study, 26 eligible participants (100% male; ages 26–53 y) were enrolled from fire stations near Sacramento, California (USA), in April–May 2025. Clinical measures from full-mouth periodontal examinations (probing depth, gingival margin position, clinical attachment loss, bleeding on probing) were compared between ONP use (≥15 in the past 30 d, no more than minimal use of other tobacco, n = 12) and tobacco nonuse (n = 14) groups. We conducted 16S rRNA gene amplicon sequencing of saliva samples to determine genus-level profiles using differential abundance testing and Bray–Curtis beta diversity analyses.
Results:
Clinical periodontal measures were numerically worse with ONP use, including a higher percentage of sites with ≥3 mm clinical attachment loss (P = 0.009). Across all samples, approximately 130 genera were detected (Streptococcus most abundant). ONP use (vs. tobacco nonuse) was associated with higher abundance of Comamonas, Alloscardovia, F0058, and Dialister and lower abundance of the [Eubacterium] sulci group, Aminipila, Mannheimia, and Peptostreptococcus. Principal coordinates analysis showed substantial overlap between groups, whereas beta dispersion was lower in the ONP group (P = 0.035).
Conclusions:
In this hypothesis-generating study, ONP use was associated with worse periodontal status and salivary microbiome shifts. While these results await confirmation in future studies and additional populations, they suggest plausible oral health risks that should be incorporated into any comprehensive assessment of potential ONP harms.
Knowledge Transfer Statement:
Although preliminary, these findings provide some of the first evidence that use of oral nicotine pouches is associated with indicators of worse periodontal health. These results should encourage further research to confirm these signals and clarify potential mechanisms. Patients, providers, and product regulators should include oral health when evaluating the risk profile of this product.
Introduction
Oral nicotine pouches (ONPs) are a growing class of noncombustible nicotine products with substantial increases in marketing, sales, and public awareness this decade (Ozga et al 2025; Travis et al 2025; Felicione et al 2026). ONPs consist of small teabag-like sachets that are usually placed in the oral vestibule and held in place, releasing nicotine (Jackson et al 2023; Felicione et al 2026). While regulated as tobacco products in some jurisdictions, most brands promote ONPs as “tobacco-free,” citing the absence of tobacco leaf matter (Ling et al 2023; Felicione et al 2026). ONPs consist largely of microcrystalline cellulose, nicotine, and flavors (Jackson et al 2023) but may contain other harmful chemicals, including tobacco-specific nitrosamines (Mallock et al 2024), albeit at lower levels than found in conventional tobacco products, as noted in industry-funded studies (Azzopardi et al 2022; Back et al 2023). By providing nicotine with less toxicity than cigarettes, there is a potential role for ONPs as a less harmful alternative to combustible tobacco (Keller-Hamilton et al 2024). However, meaningful gaps in the evidence base exist (Travis et al 2025), including implications for oral health.
It is plausible that ONP use could impair oral health, yet few industry-independent studies have examined oral health and ONP use. “Mouth lesions” and “sore mouth” were among the most reported adverse experiences among adults who used ONPs in an online survey (Dowd et al 2024). One case-series study reported oral lesions near the areas where patients held ONPs (Miluna-Meldere et al 2024b). A foreign object in the mouth could induce gingival recession, as observed with conventional smokeless tobacco use (Weintraub and Burt 1987; Robertson et al 1990). While connections between ONP use and oral conditions have not been thoroughly explored, associations between e-cigarette use and periodontal status (Atuegwu et al 2019) and oral microbiome composition (Ganesan et al 2020) suggest that use of emerging noncombustible nicotine products could influence oral health.
Periodontal disease affects the tissues that support the teeth. It is increasingly understood as a polymicrobial, dysbiosis-driven inflammatory disease in which a normally symbiotic biofilm shifts toward an inflammation-adapted community, often including enrichment of anaerobic, proteolytic taxa and microbial activities that amplify host immune activation and local tissue destruction (Hajishengallis 2015; Agrafioti et al 2022). While oral biofilm alternations have been best described in the context of periodontitis, emerging evidence suggests that similar dysbiosis–inflammation pathways may also be relevant to other disease pathways, such as oral carcinogenesis (Ting et al 2023). ONPs are held in the oral vestibule for prolonged periods and may alter local conditions (e.g., nicotine exposure, pH, salivary flow, mucosal irritation), creating selective pressures that could favor some taxa while suppressing others. Given the central role of the oral microbiome in maintaining periodontal homeostasis, it is important to evaluate whether ONP use is associated with measurable shifts in microbial community composition that could plausibly contribute to inflammation or tissue vulnerability.
Firefighters represent a population that may be particularly susceptible to ONP uptake. Growth in ONP use has been most notable among younger adults, men, and people who use other tobacco (Dai and Leventhal 2024; Palmer et al 2025). Historically, conventional oral tobacco use has been elevated among firefighters (Jitnarin et al 2015). The fire service is predominantly male, and despite strong smoke-free workplace policies, oral tobacco may be perceived as accepted among peers (Jitnarin et al 2020). ONP marketing that features workplace use, young adult models, and “tobacco-free” messaging (Ling et al 2023) may resonate with firefighters otherwise mindful of occupation-related cancer risks (Sidossis et al 2023), including oral cancer (Maloney et al 2021). The objective of this preliminary study was to explore differences in clinical periodontal status and oral microbiome composition between firefighters who used ONPs and those who used no tobacco or nicotine.
Methods
Study Design and Population
In this cross-sectional, hypothesis-generating exploratory study, investigators visited multiple fire stations within 1 fire district in the Sacramento area of California (United States) in April and May 2025. Firefighters were informed about the study in advance via posted fliers and email messages. Eligible participants were ≥18 y old and had ≥20 natural teeth. Excluded were those with a history of diabetes, a serious medical condition that would preclude a dental exam, or a current pregnancy. The University of California, San Francisco Institutional Review Board approved study procedures. Participants provided signed informed consent and received a $75 gift code redeemable with online retailers.
Participants completed a brief questionnaire to confirm eligibility and report demographic information (e.g., sex, age, race/ethnicity), height/weight, and tobacco use (Appendix). Detailed information was collected for cigarettes (lifetime, past-year, past 30-d use) and ONPs (ever, past 30 d, pouches/d). Ever and past 30-d use was collected for e-cigarettes, cigars, conventional smokeless (moist snuff, chewing tobacco, snus), hookah, and cannabis, along with household member use for all products.
After reviewing tobacco use data, investigators created 2 groups for comparison. The ONP use group required ONP use ≥15 in the past 30 d and use of other tobacco, nicotine, or cannabis no more than 3 times in the past 30 d. Inclusion in the nonuse group required no use of any tobacco, nicotine, or cannabis in the past 30 d and <100 lifetime cigarettes smoked. Participants whose tobacco use did not fit these categories were excluded from analysis. Given the exploratory nature of the study and limited existing data on potential ONP effects, sample sizes were not calculated a priori. The number of participants who fit into each use category dictated sample size, which was deemed sufficient to demonstrate feasibility and inform power calculations for future studies.
Clinical Parameters
Clinical examinations were optimized for field settings, using a portable dental chair, headlight, and single-use mouth mirrors and single-use, color-coded periodontal probes (A. Titan Instruments). A practice exercise was completed consisting of data collection on 1 young-adult volunteer to increase familiarity with the portable, single-use instruments and standardize the exam protocol. One dentist-examiner conducted all full-mouth periodontal evaluations, collecting probing depth, gingival margin position, and bleeding on probing (6 sites per tooth). Recession was defined as a gingival margin ≥2 mm below the cementoenamel junction at the facial/buccal site. Clinical attachment loss was calculated from probing depth and gingival margin position.
Statistical Analysis (Clinical Parameters)
Patient-level periodontal parameters were calculated as the mean of all probed sites and the percentage of sites above a set threshold (e.g., probing depth ≥4 mm). Nonparametric statistical hypothesis tests (Wilcoxon rank-sum test with exact P values) compared continuous periodontal parameters between ONP use and nonuse groups; chi-square tests compared any bleeding on probing and any recession (P < 0.05 considered statistically significant). Participant characteristics were summarized descriptively.
Oral Microbiome Composition
The Appendix includes additional details regarding saliva collection, processing, and analysis.
Sample Collection and Processing
Prior to clinical examinations, whole-mouth stimulated saliva was collected, then vials were secured, labeled, and placed in a portable –20°C freezer for transport (maximum 24 h) before storage at –80°C until saliva preprocessing was performed. DNA isolation was performed using the ZymoBIOMICS DNA/RNA Miniprep Kit (Zymo Research Corporation).
Amplicon Library Preparation and 16S rRNA Gene Amplicon Sequencing
The variable region 4 (V4) of the 16S rRNA gene was amplified via polymerase chain reaction (PCR), and amplicons were quantified, pooled, purified, and sequenced. Demultiplexed FASTQ files were processed to generate amplicon sequence variants (ASVs). Reads were quality filtered and trimmed, and samples with zero retained reads were excluded. Taxonomy was assigned to ASVs. An integrated phyloseq (v1.52.0) object was created from the ASV table, taxonomy, and sample metadata. Negative extraction controls and no-template controls were excluded from downstream analyses, while the mock community was retained for quality control. Relative abundances were calculated by within-sample normalization, and genus-level profiles were summarized per sample and per exposure group. Beta diversity (Bray–Curtis) was computed in phyloseq; ordination was performed by non-metric multidimensional scaling and principal coordinates analysis (PCoA), with group differences evaluated using a Wilcoxon rank-sum test. Differential genus-level relative abundance between ONP use groups was assessed using ANCOMBC2 (v2.10.1). In sensitivity analyses, we applied prevalence filtering (genera present in ≥10% of samples), excluded samples with <10,000 postchimera reads, and restricted the analysis to exclusive ONP use versus tobacco/nicotine nonuse.
Results
Study Participants
Of 36 participants who enrolled and completed study procedures, 26 met inclusion criteria for the ONP use (n = 12) or nonuse (n = 14) groups. All included participants identified as male. The 2 groups were similar in their average characteristics, including age, race/ethnicity, and body size (Table 1). No participant shared a household with someone who used combustible tobacco. No participant in the nonuse group had used any tobacco product in the past 30 d (Table 1). All 12 participants in the ONP use group used ≥15 nicotine pouches in the past 30 d, including n = 8 who used ONPs exclusively and n = 9 who used ONPs on all 30 d. Of the 4 participants who had recently used other tobacco, none used a product more than 3 in the past 30 d. All 12 participants reported using ≥4 pouches on the days they used ONPs, and n = 4 participants used ≥10 pouches/d.Table 1. Participant Characteristics and Tobacco Exposures, Nonuse and Oral Nicotine Pouch Use Groups.
| Participant Characteristic | Tobacco Nonuse (n = 14) | Oral Nicotine Pouch Use (n = 12) |
|---|---|---|
| Age, mean (range), y | 37.8 (26–53) | 34.5 (29–41) |
| Sex, % | ||
| Male | 100 | 100 |
| Race/ethnicity, % | ||
| Asian, non-Hispanic | 14.3 | 0 |
| Hispanic/Latino | 14.3 | 8.3 |
| White, non-Hispanic | 71.4 | 91.7 |
| Height, mean (SD), m | 1.82 (0.07) | 1.84 (0.06) |
| Weight, mean (SD), kg | 92.2 (14.4) | 91.9 (7.3) |
| Household exposure,a % | ||
| No tobacco | 85.7 | 66.7 |
| Smokeless only | 7.1 | 33.3 |
| Cannabis only | 7.1 | 0 |
| Cigarette use history, % | ||
| Never used | 57.1 | 8.3 |
| Former experimentationb | 42.9 | 58.3 |
| Current experimentationc | 0 | 8.3 |
| Former used | 0 | 25.0 |
| Past 30-d use of,e % | ||
| Oral nicotine pouches | 0 | 100 |
| Cigarettes | 0 | 8.3 |
| Cigars | 0 | 16.7 |
| E-cigarettes | 0 | 0 |
| Conventional smokeless | 0 | 8.3 |
| Hookah | 0 | 0 |
| Cannabis | 0 | 8.3 |
SD, standard deviation.
a
Product use by someone else who lives with a participant.
b
Smoked <100 cigarettes in their lifetime but none in the past 12 mo.
c
Smoked <100 cigarettes in their lifetime, including at least once in the past 12 mo.
d
Smoked ≥100 cigarettes in their lifetime but none in the past 12 mo.
e
Past 30-d use percentages not mutually exclusive. No product listed here (other than oral nicotine pouches) was used on >3 in the past 30 d; all participants in the oral nicotine pouch group used ≥15 pouches in the past 30 d.
Clinical Periodontal Status
All measures of periodontal status were numerically worse among participants in the ONP use group, although at this sample size, differences were statistically significant at the 0.05 threshold only for percentage of sites with ≥3 mm of clinical attachment loss (Table 2). On average, participants in the ONP use group had a higher percentage of sites with deeper pockets, ≥3 mm of clinical attachment loss, bleeding on probing, and gingival recession.Table 2. Clinical Periodontal Status, Nonuse and Oral Nicotine Pouch Use Groups.
| Participant-Levela Periodontal Status | Tobacco Nonuse (n = 14) | Oral Nicotine Pouch Use (n = 12) | P Valueb |
|---|---|---|---|
| Probing depth, mean (SD), mm | 2.0 (0.1) | 2.2 (0.4) | 0.33 |
| Probing depth, % of sites ≥4 mm | 1.6 | 7.6 | 0.13 |
| CAL, mean (SD), mm | 0.9 (0.3) | 1.3 (0.8) | 0.08 |
| CAL, % sites ≥3 mm | 5.3 | 16.5 | <0.01 |
| BOP, % of sites | 1.4 | 3.7 | 0.44 |
| Any BOP, % of participants | 64.3 | 75.0 | 0.56 |
| Recession, % of sitesc | 6.4 | 12.6 | 0.13 |
| Any recession, % of participants | 35.7 | 66.7 | 0.12 |
Abbreviations: BOP, bleeding on probing; CAL, clinical attachment loss; SD, standard deviation.
a
Measures, such as probing depth and CAL, were calculated as the mean over all sites per participant, and then the mean of those values was calculated within each tobacco use group.
b
Wilcoxon rank-sum test (exact) for continuous variables; chi-square test for any BOP and any recession.
c
Recession calculations included only facial/buccal sites.
Salivary Microbiome Composition by Exposure Group
Across the 26 saliva samples, genus-level profiling detected approximately 130 bacterial genera, with Streptococcus consistently representing the most abundant genus across both participant groups. Per-sample stacked bar plots demonstrated substantial interindividual variation in genus-level profiles within each group, while still revealing a broadly shared set of dominant oral genera (Appendix Fig. 1).
In groupwise summaries of the top 25 genera ranked by mean relative abundance, the overall core oral community structure appeared largely similar between groups; however, differences were observed in the relative rank ordering and mean abundance of several prominent genera by group (Fig. 1). Notably, Neisseria ranked among the most abundant genera in the tobacco nonuse group, whereas it exhibited a relatively lower rank and mean abundance in the ONP use group. In contrast, Veillonella and Prevotella ranked prominently in the ONP use group alongside Streptococcus (Fig. 1). Differences were also observed among lower-abundance genera included in the top 25 summaries, such as Aggregatibacter appearing among the ONP use group top genera, while Capnocytophaga and Bergeyella appeared among the tobacco nonuse group top genera (Fig. 1).
Differentially Abundant Genera
Differential abundance testing identified 8 genera with statistically significant differences between exposure groups (Table 3). Several taxa were enriched in the ONP use group relative to tobacco nonuse, including Comamonas (mean relative abundance: 1.42 × 10−5 vs. 2.15 × 10−4; log2 fold change [FC] = 2.79, P = 0.0020), Alloscardovia (9.52 × 10−5 vs. 1.16 × 10−2; log2FC = 3.31, P = 0.0093), F0058 (1.33 × 10−4 vs. 5.28 × 10−4; log2FC = 1.78, P = 0.0083), and Dialister (1.56 × 10−4 vs. 5.17 × 10−4; log2FC = 1.53, P = 0.0231). In contrast, multiple genera were depleted in the ONP use group, including the [Eubacterium] sulci group (1.13 × 10−3 vs. 6.45 × 10−4; log2FC = −1.96, P = 0.0108), Aminipila (4.18 × 10−4 vs. 1.21 × 10−4; log2FC = −2.02, P = 0.0044), Mannheimia (2.83 × 10−3 vs. 4.28 × 10−4; log2FC = −1.86, P = 0.0139), and Peptostreptococcus (1.70 × 10−3 vs. 1.04 × 10−3; log2FC = −1.40, P = 0.0200). Overall, ONP-associated differences comprised both increases in several low-abundance genera (e.g., Comamonas, F0058, Dialister) and reductions in taxa with relatively higher baseline abundance in the tobacco nonuse group (e.g., Mannheimia and [Eubacterium] sulci group), indicating that ONP use is linked to a multitaxon shift in the oral microbial community rather than a single dominant change.Table 3. Differentially Abundant Genera by 16S rRNA Gene Amplicon Sequencing between Nonuse and Oral Nicotine Pouch Use Groups.
| Genus | Tobacco Nonuse | ONP Use | Log2FCa | P Value | Family |
|---|---|---|---|---|---|
| Comamonas | 0.000014 | 0.000215 | 2.7908 | 0.00196 | Comamonadaceae |
| Alloscardovia | 0.000095 | 0.011612 | 3.3087 | 0.00932 | Bifidobacteriaceae |
| [Eubacterium] sulci group | 0.001133 | 0.000645 | –1.9559 | 0.01077 | Anaerovoracaceae |
| F0058 | 0.000133 | 0.000528 | 1.7802 | 0.00833 | Paludibacteraceae |
| Aminipila | 0.000418 | 0.000121 | –2.0183 | 0.00445 | Anaerovoracaceae |
| Mannheimia | 0.002833 | 0.000428 | –1.8564 | 0.01389 | Pasteurellaceae |
| Peptostreptococcus | 0.001696 | 0.001038 | –1.4007 | 0.01996 | Peptostreptococcaceae |
| Dialister | 0.000156 | 0.000517 | 1.5291 | 0.02311 | Veillonellaceae |
Genera with statistically significant differences in relative abundance between the tobacco nonuse and oral nicotine pouch use groups are shown. Relative abundances were calculated by within-sample normalization and summarized at the genus level from amplicon sequence variants. A mock community control was provided for quality control reference and was not included in exposure group comparisons.
a
Log2 fold change (FC) interpretation: positive values indicate higher relative abundance in the oral nicotine pouch use group; negative values indicate lower relative abundance.
Beta Diversity and Dispersion
Bray–Curtis principal coordinates analysis (PCoA) showed overlap in community profiles between participant groups with partial separation, suggesting that any exposure-associated differences in overall community structure were modest in this sample (Fig. 2A). The first 2 axes explained 48.5% (principal component [PC] 1) and 18.8% (PC2) of the variance. Beta dispersion (distance to centroid) was significantly lower in the ONP use group (P = 0.035; Fig. 2B), indicating differences in within-group heterogeneity.
Sensitivity Analyses
Clinical findings did not differ meaningfully when restricted to n = 8 participants who used ONPs exclusively and the n = 14 participants in the tobacco nonuse group (Appendix Table 1). Likewise, Bray–Curtis PCoA (1 sample excluded for having <10,000 postchimera reads) showed substantial overlap in community composition with only partial separation, consistent with modest between-group differences in overall community structure (Appendix Fig. 2A). The first 2 ordination axes explained 49.2% (PCoA1) and 16.1% (PCoA2) of the variance. Beta dispersion (distance to group centroid) was significantly lower in the exclusive ONP use group (P = 0.0085; Appendix Fig. 2B), indicating reduced within-group compositional variability relative to tobacco nonuse.
Differential abundance testing identified 4 genera with statistically significant differences (Benjamini-Hochberg-adjusted P < 0.05; Appendix Fig. 2C). Three taxa were enriched in the exclusive ONP use group (mean relative abundance in tobacco nonuse vs. exclusive ONP use: Comamonas 1.42 × 10−5 vs. 1.94 × 10−4, log2FC = 2.30, P = 0.0279; Dialister 1.56 × 10−4 vs. 7.21 × 10−4, log2FC = 1.71, P = 0.0317; and F0058 1.33 × 10−4 vs. 3.56 × 10−4, log2FC = 1.46, P = 0.0414). In contrast, Aminipila was depleted in the exclusive ONP use group (4.18 × 10−4 vs. 2.07 × 10−4, log2FC = −1.85, P = 0.0201).
Discussion
In this preliminary investigation, multiple indicators of periodontal health, including clinical and microbiome parameters, suggested worse periodontal status among participants who used ONPs compared to participants who used no tobacco or nicotine. While these findings are notable, a cautious interpretation is advised given the cross-sectional design, modest sample size, and need for testing in other populations. Nonetheless, the rising popularity of ONPs, societal burden of oral diseases, and relative lack of existing evidence all provide reasons for this hypothesis-generating investigation and for future research to validate these results in additional study populations.
In the present study, participants who used ONPs had greater clinical attachment loss, bleeding on probing, and gingival recession, though only attachment loss was statistically significant. Our exploratory data also suggest an ONP-related restructuring of the oral microbiome. In descriptive summaries, Neisseria, a periodontal health indicator (Yamashita and Takeshita 2017; Fukuda et al 2024), was lower in rank and mean abundance among participants who used ONPs. Differential abundance testing identified 8 genera with statistically significant differences between groups and lower within-group beta dispersion within the ONP use group, suggesting more homogeneous community profiles. These taxa-level signals should be interpreted cautiously, but the microbiome and clinical findings together provide convergent evidence to motivate further research, including integrated analyses of proteomic, microbiome, and inflammatory pathways that may link ONP exposure to periodontal disease.
Although genus-level summaries and differential abundance testing identified several taxa-level differences, Bray–Curtis PCoA showed substantial overlap between groups, suggesting that ONP-associated differences in overall salivary community structure were modest. Notably, beta dispersion was lower with ONP use, consistent with convergence toward more similar community profiles, which could reflect repeated, localized exposure conditions acting as an “environmental filter” on the oral microbiome. Because dispersion can influence apparent group separation in ordination-based summaries, these community-level findings should be interpreted cautiously and validated in larger, longitudinal studies with site-specific sampling (e.g., mucosa adjacent to pouch placement and subgingival plaque), which may better capture localized exposure effects than whole saliva.
Microbiome shifts are relevant because periodontal disease is a dysbiosis-driven condition in which community structure changes, rather than single pathogens, can influence host responses and clinical outcomes (Hajishengallis 2015). Even modest compositional changes, particularly reductions in health-associated commensals (like Neisseria) or enrichment of inflammation-tolerant taxa, could have downstream implications for biofilm resilience and periodontal stability (Hajishengallis 2015; Cugini et al 2021). Periodontal pathobionts may have additional disease implications, plausibly as potential contributors to protumorigenic signaling and immune modulation in oral squamous cell carcinoma (Lafuente Ibáñez de Mendoza et al 2020; McIlvanna et al 2021).
It should be emphasized that the firefighters in this study represent a largely healthy, relatively young, community-dwelling sample that, as a group, presented without extensive periodontal disease. Exclusively male, this sample is not reflective of the general population. Speculatively, occupational stress or exposures specific to firefighters might influence nicotine use and microbiome status. However, the firefighter sample offers some advantages over clinical patient samples, as patients may have more advanced periodontal disease than the general population. Thus, the present study might be considered a window into earlier stages of the disease process.
The present findings align with the existing research that suggests ONPs might exert adverse oral health effects (Miluna-Meldere et al 2024a, 2024b; Alkhatib 2025). Gingival recession has previously been related to use of oral tobacco (Weintraub and Burt 1987; Robertson et al 1990). Speculatively, consistent application of ONPs directly against the gingival tissue could contribute mechanically to recession. Possible nicotine effects, pH changes, or mucosal irritation are other speculative mechanisms. Future research should explore localized recession in areas where ONPs are typically held, as highlighted in case reports (Alkhatib 2025). Notably, the increased bleeding on probing observed in the ONP use group contrasts with the well-documented decreased vascular density and blood vessel narrowing associated with cigarette smoking (Bergström et al 1988). This contrast merits further investigation, especially given evidence connecting e-cigarette use to gum bleeding (Silveira et al 2022). Indeed, several studies connect e-cigarette use to adverse oral health outcomes, such as tissue irritation, gingivitis, xerostomia, and oral microflora alterations (Atuegwu et al 2019; Ganesan et al 2020; Silveira et al 2022; Panariello et al 2025; Chaffee 2025). While different from ONPs in composition and mode of nicotine delivery, these e-cigarette studies provide some indication that use of noncombustible nicotine products, like ONPs, could affect the oral cavity.
Oral health is one consideration within a broader question of whether ONPs offer a public health benefit as a potentially less harmful form of nicotine delivery compared to combustible cigarettes. ONP use prevalence is currently low but has grown rapidly (Travis et al 2025; Felicione et al 2026). The tobacco industry has an emerging research interest in oral health, with several industry-connected studies framing ONPs as less damaging than other tobacco products for tooth stain, gingivitis, mouth irritation, and other outcomes (La Rosa et al 2025; Liu et al 2025). While it is likely that ONPs are less dangerous than conventional tobacco products, there is a need to understand the absolute health impacts of ONP use. Any potential benefits as an alternative to other tobacco should be weighed against potential ONP-related health risks and appeal to youth (Travis et al 2025).
In this study, some individuals in the ONP group had recently used other tobacco products, potentially affecting their periodontal condition. However, exclusion of these individuals from sensitivity analyses of clinical data did not meaningfully affect the results, and key microbiome signals (including Comamonas, Dialister, F0058, Aminipila, and reduced beta dispersion) persisted. Dual or poly-use of multiple tobacco and/or nicotine products is increasingly common (Hirschtick et al 2021), making it difficult to enroll study populations defined by exclusive use of 1 product. Statistical adjustment for other product use, as well as other plausible confounding variables, can be unreliable in small samples but should be considered for larger, confirmatory studies.
Several additional study limitations deserve consideration. Prominently, microbiome findings must be validated in additional, independent cohorts. As a preliminary study designed in part to generate hypotheses and inform future protocols, statistical power was limited to detect some potentially meaningful differences. Results were not adjusted for potential confounding factors, such as oral hygiene practices, alcohol use, or past tobacco smoking. Thus, the potential role of residual confounding should be considered. However, the overall level of past cigarette smoking in the ONP group was low, and associations persisted when analyses were restricted to participants who used ONPs exclusively. Tobacco use was self-reported; although self-reported tobacco status is generally valid (Patrick et al 1994), biomarker verification would strengthen future work. Further studies are warranted that consider specific aspects of ONP products, including nicotine concentration and flavors.
In summary, this preliminary study provides new clinical and microbial evidence of possible associations between the use of oral nicotine pouches and adverse oral health—a signal worthy of further examination in larger samples. To inform appropriate product regulation and patient communication, the potential for adverse oral health effects from nicotine pouch use should be examined further.
Author Contributions
B.W. Chaffee, contributed to conception and design, data acquisition, analysis, and interpretation, drafted the manuscript; K. Lieuw, contributed to acquisition, analysis, and interpretation, critically revised the manuscript; H. Zhang, contributed to analysis and interpretation, critically revised the manuscript; E.T. Couch, contributed to conception and design, data acquisition and interpretation, critically revised the manuscript; Y.J. Jiang, contributed to analysis, critically revised the manuscript; F. Momen-Heravi, contributed to conception and design, data analysis and interpretation, drafted the manuscript. All authors gave their final approval and agreed to be accountable for all aspects of the work.
Acknowledgments
The authors thank Andrew Nguyen of UCSF for assistance in data collection and sample processing, Lisa Berens of UCSF for providing portable dental equipment, and the UCSF Benioff Center for Microbiome Medicine Microbial Genomics CoLab Plug-in for its assistance with data generation.
Declaration of Conflicting Interests
The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This work was supported by funding from the US National Institutes of Health (R01DE032433 and U01DE033351 to F. Momen-Heravi). Support was received from the University of California, San Francisco (UCSF) Lee Hysan Fund.